INTERACTION OF BIFIDOBACTERIA WITH THP1 MONOCYTIC CELL LINE

DIAZ, MARIÁNGELES; ROLNY, IVANNA; MINNAARD, JESSICA AND PÉREZ, PABLO F.

Tucuman

Simposio; IV International Symposium on Lactic Acid Bacteria. Food, Health and Applications.; 2013

Interaction between bifidobacteria and host´s cells is relevant for the health promoting effects and studies with professional phagocytic cells could contribute to the understanding of the mechanisms involved in the probiotic effect. The present work aimed to evaluated the interaction of bifidobacteria with monocytic cells from human origin (THP1 cells). Bifidobacterium bifidum 5310 and B. adolescentis 5317 were cultured (48 hs; 37°C) in MRS broth in anaerobic conditions. Cultures were centrifuged, washed and suspended in DMEM medium. Bacteria were labeled with FITC. THP1 cells (1 x 106 cell/ml) were differentiated with phorbol myristate acetate (PMA) 200 nm in DMEM 10% fetal bovine serum for 3 days at 37°C in 5% CO2 atmosphere.. Coincubations (THP1 cells-bacteria) were performed for 1 h at 37°C at different multiplicities of infection (MOI). The effect of the presence of Bacillus cereus B10502 (MOI 25 or 50 for 30 min) was also studied. Association and invasion were evaluated by flow cytometry (FL1+ cells), epifluorescence microscopy and plate counts in MRS agar. Necrosis was assessed by flow cytometry after propidium iodide staining (FL2+ cells). Internalization was evaluated after quenching of exocellular bacteria with trypan blue. Bacteria were found as large clusters around the monocytes. This was particularly evident for strain CIDCA 5310. Flow cytometry analysis showed 92.3 ± 4.8 % of cells associated to strain CIDCA 5310 (FL1 + events) irrespectively of the MOI assayed. These findings suggest that monocyte-bacteria interaction is saturated at low MOI. In contrast, strain CIDCA 5317 showed a dose-response behavior; i. e. 12.6 ± 3.8 %; 30.3 ± 6.0 % and 61.2 ± 17.6 % for MOI 25, 50 and 100 respectively. After quenching with trypan blue, percentage of FL1 + cells ranged from 1 to 5 % thus suggesting that high number of bacteria remained exocellular. MOI of 50 and higher of strain CIDCA 5310 lead to around 70 % necrosis (FL2+ cells) whereas values remains similar to controls (20 %) for strain CIDCA 5317. When monocytes were incubated with bifidobacteria and strain B10502 of Bacillus cereus a 2 to 3 fold increase in the percentage of cells associated to bifidobacteria was found. Results demonstrated that bifidobacteria interact with phagocytic cells in a strain-dependent manner. These findings could be related to differences in surface properties already reported for the strains under study and could be relevant for the understanding of the immunomodulating effects of these potentially probiotic strains.